[GnRH-a combined fertility-sparing re-treatment in women with endometrial carcinoma or atypical endomertial hyperplasia who failed to oral progestin therapy].

Objective: To analyze the clinical efficacy and pregnancy outcomes of gonadotropin-releasing hormone agonist (GnRH-a) based fertility-sparing re-treatment in women with endometrial carcinoma (EC) and atypical endometrial hyperplasia (AEH) who failed with oral progestin therapy. Methods: Forty cases with EC or AEH who failed to respond to oral progestin were included from January 2012 to December 2020 at Peking Union Medical College Hospital. Combination of GnRH-a with levonorgestrel-releasing intrauterine system (group GLI: a subcutaneous injection of GnRH-a every 4 weeks and LNG-IUS insertion constantly) or the combination of GnRH-a with aromatase inhibitor (group GAI: a subcutaneous injection of GnRH-a every 4 weeks and oral letrozole 2.5 mg, daily) were used for these patients. Histological evaluation were performed at the end of each course (every 3-4 months) by hysteroscopy and curettage. After the complete remission (CR), all patients were followed up regularly. Results: (1) Clinical characteristics:among the 40 patients with EC or AEH, the median age at diagnosis was 31 years (range: 22-40 years) and the median body mass index was 24.7 kg/m2 (range: 18.9-39.5 kg/m2). (2) Efficacy of fertility-sparing re-treatment: 37 (92%, 37/40) patients achieved CR, 6 (6/7) in AEH and 31 (94%, 31/33) in EC patients. The CR rate was 93% (26/28) and 11/12 in group GLI and GAI, respectively. The median time to CR was 5 months (range: 3-12 months). At the end of the first therapy course, the CR rates in AEH and EC were 5/7 and 42% (14/33), at the second course, the CR rates were 6/7 and 82% (27/33), respectively. (3) Recurrence: after 25 months of median follow-up duration (range: 10-75 months), 8 (22%, 8/37) women developed recurrence, 1/6 in AEH and 7 (23%, 7/31) in EC patients, with the median recurrence time of 18 months (range: 9-26 months). Among them, two cases who had completed childbirth chose to receive hysterectomy directly. Six patients met the criteria of fertility-preserving therapy and received conservative treatment again and 5 (5/6) of them achieved CR. (4) Pregnancy: of the 37 patients with CR, 33 desired to conceive. Ten women attempted to get pregnancy spontaneously and 23 cases with assisted reproductive technology. Fourteen (42%, 14/33) patients became pregnant, including 9 (27%, 9/33) live births, 3 (9%, 3/33) missed abortions, and 2 (6%, 2/33) miscarriages at the second trimester. Conclusions: GnRH-a based fertility-sparing re-treatment in AEH or EC patients who failed with oral progestin therapy achieved good treatment effect and reproductive outcomes. It is an encouraging alternative regime for patients who failed with oral progestin therapy.

Comparative analysis of microbial community in the whole body and midgut from fully engorged and unfed female adult Melophagus ovinus.

Melophagus ovinus is a type of ectoparasite infesting sheep. Data regarding the comprehensive bacterial community associated with the whole body and midgut of M. ovinus under different engorged statuses are required. Melophagus ovinus were collected from the city of Jiuquan, China. Bacterial DNA was extracted from the whole body and midgut of fully engorged female adults, or newly hatched and unfed adult female M. ovinus. The 16S rRNA gene V3-V4 hypervariable regions were sequenced using the IonS5™XL platform (Thermo Fisher Scientific, Waltham, MA, U.S.A.). The whole body bacterial diversity of the newly hatched, unfed adult females was greater compared with that of the other three samples. Proteobacteria was the dominant bacterial phylum in all of the samples. Of the 42 total bacterial genera present in all of the experimental samples, Arsenophonus, Bartonella and Wolbachia were the dominant genera. The relative abundance of Arsenophonus in midgut was greater than that in the whole body. The relative abundance of Bartonella in fully engorged adults was far greater than those in newly hatched, unfed adults. The relative abundance of Wolbachia was highest in the whole body of newly hatched, unfed adults. Seventeen bacterial species were identified in all experimental samples. Bartonella chomelii, Streptococcus hyointestinalis and Escherichia coli were the first species reported in M. ovinus.

[Role of short-term starvation in alleviating hepatic ischemia-reperfusion injury in mice and possible mechanism of action].

Objective: To investigate the role of short-term starvation (STS) in alleviating hepatic ischemia-reperfusion injury in mice and possible mechanism of action. Methods: Wild-type male C57BL/6 mice aged 8 weeks were randomly divided into 75% hepatic ischemia-reperfusion injury group (IR group), STS+75% hepatic ischemia-reperfusion injury group (STS group), and sirtinol+STS+75% hepatic ischemia-reperfusion injury group (SIR group), using a random number table, and sham-operation groups (IR-Sham group, STS-Sham group, and SIR-Sham group) were also established. The serum levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were measured, and the histomorphological changes of the liver were observed, as well as the expression of Sirt1, LC3B, and P62 proteins in liver tissue and the results of LC3B fluorescence staining. An analysis of variance was used for comparison of data between multiple groups, and the t-test was used for comparison of data between two groups. Results: Compared with the IR group, the STS group had significant reductions in the serum levels of ALT (3 152.7 ± 735.6 U/L vs 8 414.2 ± 1 052.2 U/L, P < 0.01) and AST (3 577.0 ± 714.0 U/L vs 10 845.8 ± 1 145.7 U/L, P < 0.01) and significant alleviation of liver pathological injury (Suzuki score: 1.50±0.55 vs 3.50±0.55, P < 0.01). Compared with the STS group, the SIR group had significant increases in the serum levels of ALT (7 002.7 ± 1 485.2 U/L vs 3 152.7 ± 735.6 U/L, P < 0.01) and AST (8 980.7 ± 1 739.1 U/L vs 3 577.0 ± 714.0 U/L, P < 0.01) and significant exacerbation of liver pathological injury (Suzuki score: 3.33 ± 0.52 vs 1.50 ± 0.55, P < 0.01). Compared with the IR group and the IR-Sham group, the STS group and the STS-Sham group had significant increases in the mRNA and protein expression of Sirt1 and the protein expression of LC3B and a significant reduction in the protein expression of P62, as well as a significant increase in the percentage of LC3B-positive cells in liver tissue (22.83% ± 5.19% / 22.17% ± 4.83% vs 10.16% ± 3.06% / 10.83% ± 1.94%, both P < 0.01). Compared with the STS group and the STS-Sham group, the SIR group and the SIR-Sham group had significant reductions in the expression of Sirt1 and LC3B proteins and a significant increase in the expression of P62 protein, as well as a significant reduction in the percentage of LC3B-positive cells in liver tissue (11.83% ± 9.24% / 14.67% ± 4.68% vs 22.83% ± 5.19% / 22.17% ± 4.83%, both P < 0.01). Conclusion: STS can effectively alleviate hepatic ischemia-reperfusion injury, and its protective effect may be associated with increasing the expression of Sirt1, inducing and promoting hepatocyte autophagy, and reducing hepatocyte death.

[Relationship between the Change Rules of Volatile Organic Compounds in Rat Muscle and Postmortem Interval].

OBJECTIVES: To explore the relationship between the change rules of volatile organic compounds （VOCs） in rat muscle and postmortem interval （PMI）. METHODS: A total of 120 healthy rats were divided randomly into 12 groups （10 for each group）. After the rats were sacrificed by cervical dislocation, the bodies were kept at （25±1） ℃. Rat muscle samples were separately obtained at 12 PMI points, including 0, 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9 and 10 d. The VOCs in rat muscles were collected, detected and analyzed by headspace solid-phase microextraction （HS-SPME） coupled to gas chromatography-mass spectrometer （GC-MS）. RESULTS: In total, 15 species of VOCs were identified, including 9 aromatic compounds, 3 sulfur compounds, 2 aliphatic acids and 1 heterocyclic compound. The species of VOCs increased with PMI： no species were detected within 1 day, 3 species were detected on day 2, 9 on day 3, 11 on day 4, 14 from day 5 to 7, and 15 from day 8 to 10. Total peak area of 15 species of VOCs was significantly correlated to PMI （adjusted R²=0.15-0.96）： the regression function was y=-17.05 x²+ 164.36 x-246.36 （adjusted R²=0.96） from day 2 to 5, and y=2.24 x+101.13 （adjusted R²=0.97） from day 6 to 10. CONCLUSIONS: The change rules of VOCs in rat muscle are helpful for PMI estimation.

Effects of carbomer eye drops in combination with orthokeratology lens in treating adolescent myopia.

To investigate the effects of carbomer eye drops (CED) during long-time wearing of overnight orthokeratology lens of adolescents with myopia, 260 teenagers with myopia treated in the Henan Provincial People’s Hospital from June 2012 to August 2014 and followed-up for more than 2 years were enrolled. All the patients underwent regular fitting of orthokeratology lens. They were divided into a CED (Vidisic) group (130 cases, 260 eyes treated with CED) and rewetting drops (RD) (Baushe and Lomb) group (130 cases, 260 eyes treated with RD). The effects in the two groups were observed. The incidence of corneal epithelial defects one day, one week and one month after treatment of the CED group was lower than that of the RD group, and the difference was statistically significant (P less than 0.05); the tear break up time (TBUT) of the CED group was higher than that of the RD group at different time points, and the difference had statistical significance (P less than 0.05); the difference of the value of Schirmer I test between the two groups had no statistical significance (P>0.05). It is concluded that carbomer eye drops can stabilize tear film and protect and repair corneal epithelium during the wearing of orthokeratology lens.

Hyperglycemia and liver ischemia reperfusion injury: a role for the advanced glycation endproduct and its receptor pathway.

Although pretransplant diabetes is a risk factor for mortality post-liver transplant, the underlying mechanism has not been fully defined. In a murine liver partial warm ischemia model, we addressed the question of how diabetes/hyperglycemia impacted tissue inflammatory injuries against ischemia reperfusion (IR), focusing on the advanced glycation endproduct (AGE) and its receptor (RAGE) pathway. Our results showed that hepatocellular injury was exacerbated in streptozotocin-induced diabetic mice against IR, in association with hyper-inflammatory immune activation in livers. Serum levels of AGEs, but not HMGB1, were increased in diabetic mice in response to liver IR. Both RAGE antagonist peptides and small interfering RNA alleviated liver injuries and inhibited inflammatory immune activation against IR in diabetic, but not normal, mice. Kupffer cells (KCs)/macrophages, but not hepatocytes, from diabetic mice expressed significantly higher levels of RAGE, leading to their hyper-inflammatory responsiveness to both TLR ligands and AGEs. In vitro, hyperglycemia increased macrophage RAGE expression and enhanced their TLR responses. Our results demonstrated that activation of the AGE-RAGE signaling pathway in KCs was responsible for hyper-inflammatory immune responses and exacerbated hepatocellular injuries in diabetic/hyperglycemic hosts against liver IR.

Construction and characterization of recombinant adenovirus carrying a mouse TIGIT-GFP gene.

Recombinant adenovirus vector systems have been used extensively in protein research and gene therapy. However, the construction and characterization of recombinant adenovirus is a tedious and time-consuming process. TIGIT is a recently discovered immunosuppressive molecule that plays an important role in maintaining immunological balance. The construction of recombinant adenovirus mediating TIGIT expression must be simplified to facilitate its use in the study of TIGIT. In this study, the TIGIT gene was combined with green fluorescent protein (GFP); the TIGIT-GFP gene was inserted into a gateway plasmid to construct a TIGIT-GFP adenovirus. HEK 293A cells were infected with the adenovirus, which was then purified and subjected to virus titering. TIGIT-GFP adenovirus was characterized by flow cytometry and immunofluorescence, and its expression in mouse liver was detected by infection through caudal vein injection. The results showed the successful construction of the TIGIT-GFP adenovirus (5 x 10(10) PFU/mL). Co-expression of TIGIT and GFP was identified in 293A and liver cells; synthesis and positioning of TIGIT-GFP was viewed under a fluorescence microscope. TIGIT-GFP was highly expressed on liver cells 1 day (25.53%) after infection and faded 3 days (11.36%) after injection. In conclusion, the fusion of TIGIT with GFP allows easy, rapid, and uncomplicated detection of TIGIT translation. The construction of a TIGIT-GFP adenovirus, mediating TIGIT expression in vitro and in vivo, lays the foundation for further research into TIGIT function and gene therapy. Moreover, the TIGIT-GFP adenovirus is a helpful tool for studying other proteins (which could replace the TIGIT gene).

[Clinical characteristics of 42 children with focal atrial tachycardia originated from the right atrial appendage].

Objective: To investigate the feature and treatment of atrial tachycardia (AT) originated from right atrial appendage (RAA) in children. Methods: The data of 42 children with AT originated from RAA, who were admitted the First Hospital of Tsinghua University from January 2010 to September 2022 were analyzed retrospectively.The clinical characteristics, treatment and efficacy were analyzed. The children were divided into tachycardia cardiomyopathy group and normal cardiac function group. The differences in the ablation age and the heart rate during AT between two groups were compared by independent sample t-test. Results: Among 42 children, there were 20 males and 22 females. The age of onset was 2.7 (0.6, 5.1) years. Their age at radiofrequency ablation was (6.5±3.6) years, and the weight was (23.4±10.0) kg. Thirty-two children (76%) had sustained AT. The incidence of tachycardia cardiomyopathy was 43% (18/42). Compared to that of the normal cardiac function group, the ablation age and the heart rate at atrial tachycardia of the tachycardia cardiomyopathy group were higher ((8.1±3.8) vs. (5.3±3.1) years, t=-2.63, P=0.012; (173±41) vs. (150±30) beats per minute, t=-2.05, P=0.047. Thirty-eight children (90%) responded poorly to two or more antiarrhythmic drugs. The immediate success rate of radiofrequency ablation (RFCA) was 57% (24/42), and the AT recurrence rate was 17% (4/24). Twenty-two children underwent RAA resection, and their AT were all converted to sinus rhythm after the surgery. During the RAA resection, 10 cases of right atrial appendage aneurysm were found, 9/18 of which failed the RFCA. Conclusions: The AT originated from the RAA in children tend to present with sustained AT, respond poorly to antiarrhythmic drugs, and has a low success rate of RFCA as well as high recurrence rate. Resection of the RAA is a safe and effective complementary treatment.

DNA methylation status in tissues of sheep clones.

Genomic imprinting and DNA methylation play an important role in mammalian development. Many cloned animals showed heterogeneous DNA methylation profiles. However, there are fewer reports in cloned lambs because of a lack of genomic imprinting information. In this study, we investigated DNA methylation patterns in CpG islands and differentially methylated regions of putative imprinted gene Peg10 and imprinted genes Dlk1, Igf2R and H19 in cloned lambs. Five organs from two cloned lambs died shortly after birth and two normal controls were investigated. We observed normal DNA methylation profiles in cloned lambs. The imprinted genes Dlk1, Igf2R and H19 in livers, kidneys, hearts, muscles and lungs of the two cloned lambs exhibited relatively normal DNA methylation, except for Peg10 showing some differences between controls and cloned lambs. Our results indicate that somatic cell nuclear transfer-produced sheep exhibited relatively normal DNA methylation pattern and experienced normal DNA methylation reprogramming at imprinted loci.

[Effectiveness investigation on left bundle branch area pacing in 10 infants and toddlers].

Objective: To explore the feasibility, safety and effectiveness of left bundle branch area pacing (LBBAP) in children aged ≤3 years. Methods: A total of 10 children aged ≤3 years who were diagnosed with brady arrhythmia in the First Hospital of Tsinghua University from September 2020 to September 2021 were retrospectively analyzed. All the children met the indication of permanent pacemaker implantation and underwent LBBAP successfully. The intraoperative data (pacing parameters, electrocardiogram and radiographic imaging), cardiac ultrasound data and clinical data during regular postoperative follow-up were recorded. The preoperative and postoperative data were compared using matched samples t test. Results: Ten children (aged (1.6±0.7) years with weight of (10.3±2.5) kg) underwent LBBAP successfully. The QRS wave duration on the postoperative electrocardiogram was (100±9) ms, and the percentage of ventricular pacing was (97±7)%. The postoperative follow-up period was 6 (6, 12) months. At 1 week after operation, the left ventricular end-diastolic diameter Z scores in these children reduced significantly compared with those before operation (1.3±0.6 vs. 3.6±1.1, t=9.37, P<0.001). During the follow-up period, cardiac function was normal and the last left ventricular ejection fraction was (66±4)% in all children. At the last follow-up, the pacing threshold of the 10 children was smaller than 1.0 V and was acceptable. Compared with the intraoperative baseline values, the pacing threshold was slightly higher ((0.8±0.1) vs. (0.5±0.1) V, t=-5.27, P=0.001). However, no significant difference was found regarding sensing threshold ((16±5) vs. (14±4) mV, t=-0.83, P=0.426) and impedance ((584±88) vs. (652±86) Ω, t=2.26, P=0.050). During follow-up, no electrode related complications were recorded. Conclusions: LBBAP is safe and effective for infants and toddlers. Narrow QRS pacing with stable pacing parameters and normal cardiac function could be achieved postoperatively.

Fabrication and characterization of elastomeric polyester/carbon nanotubes nanocomposites for biomedical application.

A novel biodegradable polymer elastomer nanocomposite composing of poly(1,8-octanediol-citrate) (POC) polymer matrix and carbon nanotubes (CNTs) additive was successfully fabricated and systematically investigated using Fourier transform infrared (FT-IR), X-ray diffractometer (XRD), differential scanning calorimetry (DSC), tensile test, incubation and cytotoxicity tests. It was found that the addition of CNTs in POC elastomer did not result in any noticeable change in its chemical structure and the amorphous state. However, the tensile strength and elongation at break were greatly improved by the addition of CNTs in POC polymer matrix. It revealed that the swelling ratio and percentage of weight loss of POC/CNTs nanocomposite were lower, compared with the pure POC material. Moreover, the adsorption amount of bovine serum albumin (BSA) increased with an increase of the CNTs mass content in POC matrix revealing the enhanced hydrophilicity of POC/CNTs nanocomposites contributed by the carboxyl of the CNTs. Additionally, the cytotoxicity tests with L929 cell line revealed that the experimental POC/CNTs nanocomposites possessed good in vitro biocompatibility.

Human papillomaviruses in oral carcinoma and oral potentially malignant disorders: a systematic review.

OBJECTIVES: Human papillomavirus (HPV) in oral carcinoma (OSCC) and potentially malignant disorders (OPMD) is controversial. The primary aim was to calculate pooled risk estimates for the association of HPV with OSCC and OPMD when compared with healthy oral mucosa as controls. We also examined the effects of sampling techniques on HPV detection rates. METHODS: Systematic review was performed using PubMed (January 1966-September 2010) and EMBASE (January 1990-September 2010). Eligible studies included randomized controlled, cohort and cross-sectional studies. Pooled data were analysed by calculating odds ratios, using a random effects model. Risk of bias was based on characteristics of study group, appropriateness of the control group and prospective design. RESULTS: Of the 1121 publications identified, 39 cross-sectional studies met the inclusion criteria. Collectively, 1885 cases and 2248 controls of OSCC and 956 cases and 675 controls of OPMD were available for analysis. Significant association was found between pooled HPV-DNA detection and OSCC (OR = 3.98; 95% CI: 2.62-6.02) and even for HPV16 only (OR = 3.86; 95% CI: 2.16-6.86). HPV was also associated with OPMD (OR = 3.87; 95% CI: 2.87-5.21). In a subgroup analysis of OPMD, HPV was also associated with oral leukoplakia (OR = 4.03; 95% CI: 2.34-6.92), oral lichen planus (OR = 5.12; 95% CI: 2.40-10.93), and epithelial dysplasia (OR = 5.10; 95% CI: 2.03-12.80). CONCLUSIONS: The results suggest a potentially important causal association between HPV and OSCC and OPMD.

Propofol suppresses proliferation, migration and invasion of gastric cancer cells via regulating miR-29/MMP-2 axis.

The article "Propofol suppresses proliferation, migration and invasion of gastric cancer cells via regulating miR-29/MMP-2 axis" by Y.-J. Ni, J. Lu, H.-M. Zhou, published in Eur Rev Med Pharmacol Sci 2019; 23(19): 8606-8615 has been withdrawn.

Propofol suppresses proliferation, migration and invasion of gastric cancer cells via regulating miR-29/MMP-2 axis.

OBJECTIVE: Propofol (2,6-diisopropylphenol) is a commonly used intravenous anesthetic agent. Previous studies suggested that propofol might act as anti-tumor drug in various cancers, including gastric cancer. However, the underlying mechanism is still largely unknown. MATERIALS AND METHODS: 1, 5, 10 and 20 µg/ml of propofol were used to treat gastric cancer cell MKN45 for 24, 48 or 72 hours. MTT assay was used to detect the proliferation. Transwell assay was employed to measure the invasion and migration with or without matrigel. The expression of miR-29a, 29b and 29c was assessed by quantitative real time polymerase chain reaction (qRT-PCR). Luciferase assay was introduced to confirm the relationship between miR-29 family member and MMP-2. Western blot was adopted to measure the expression of MMP-2 protein. RESULTS: The proliferation, migration and invasion of gastric cancer cell MKN45 were gradually decreased after propofol treatment in time- and dose- dependent manners. MiR-29a, b and c were downregulated in MKN45 cells compared with normal gastric mucosa epithelial cell GES-1 and upregulated by propofol. Inhibition of miR-29a, b or c promoted cell proliferation, migration and invasion of MKN45 cells under propofol treatment. MMP-2 was a target and regulated by miR-29 family and propofol. MMP-2 silencing reversed the stimulative effects of miR-29 inhibitor. CONCLUSIONS: Propofol inhibited cell proliferation, migration and invasion by upregulating miR-29a, miR-29b and miR-29c and downregulating MMP-2.

[Results of microscopic examinations and blood slide re-examinations of malaria elimination evaluation in ten prefectures of Yunnan Province].

OBJECTIVE: To understand the laboratory technicians' abilities in blood slide making and reading in 10 prefectures of Yunnan Province which have passed the provincial malaria elimination evaluation, so as to provide the evidence for improving the malaria elimination surveillance and parasite examination. METHODS: Thirty negative blood slides were randomly sampled to evaluate coating, dyeing and clean quality and reading results, and 4 laboratory technicians were sampled to evaluate their reading abilities from each prefecture level and its 2 subordinate counties (districts) respectively, and then the results were analyzed. RESULTS: A total of 869 negative blood samples were evaluated. The coincidence rate was 100%. The proportions of good coating, dyeing and clean quality were 96.09%, 91.71% and 96.89%, respectively. Totally 576 blood slides were used to evaluate the reading ability. The number of correct reading was 505, and the correct rate was 87.67%. Among them, the Plasmodium vivax correct reading rate was 87.76%, the P. falciparum correct reading rate was 87.50%, and the correct reading rate of mixed infections was 47.62%. The laboratory technicians' ability to the mixed infections was significantly lower than the ability to the others (χ2 = 37.169, P < 0.05), however, in the laboratory technicians' abilities, there was no significant difference among the center (s) for disease control and prevention, general hospitals and township hospitals (χ2 = 2.782, P > 0.05), and the prefecture, county and township levels (χ2 = 0.358, P > 0.05) . CONCLUSIONS: The 10 prefectures have passed the provincial evaluation in blood slide making and microscopic examination skill indicators requested, but the medical and public health institutions at all levels still should further improve their laboratory technicians' abilities in blood slide making and microscopic examination skills.

Macrophages Promote Growth of Squamous Cancer Independent of T cells.

Oral cancers, primarily squamous cell carcinomas (SCCs), progress either slowly or aggressively. Here we assessed the role of macrophages in SCC behavior. We used mouse SCC cells derived from tumors harboring a KrasG12D activation mutation and Smad4 deletion in keratin 15-positive stem cells and a human oral SCC cell line, FaDu, which has NRAS amplification and SMAD4 deletion. SCC cells were transplanted into immune-compromised or immune-competent (syngeneic) recipients. After tumors were established, we used clodronate liposomes to ablate macrophages. We found that the number of tumor-associated macrophages (TAMs) was not affected by the presence of T cells but differed considerably among tumors derived from different SCC lines. Clodronate significantly reduced TAMs and splenic macrophages, resulting in reduced SCC volumes. Tumors with clodronate treatment did not show decreased proliferation but did exhibit increased apoptosis and reduced vascular density. FLIP (Fas-associated via death domain-like interleukin 1ß-converting enzyme inhibitory protein), an apoptosis inhibitor abundantly produced in tumor cells and TAMs, was reduced in tumor cells of clodronate-treated mice. Reduced FLIP levels correlated with reductions in phosphorylated nuclear NFκB p65 and NFκB inhibitor attenuated FLIP protein levels in SCC cells. Furthermore, TGFß1 serum levels and pSmad3 were reduced in clodronate-treated mice, but their reductions were insufficient to reverse epithelial-mesenchymal transition or TGFß-mediated angiogenesis in endothelial cells. Consequently, metastasis was not significantly reduced by macrophage reduction. However, reduced pSmad3 correlated with reduction of its transcriptional target, vascular endothelial growth factor A, in clodronate-treated tumor cells, which correlated with reduced vascular density in clodronate-treated tumors. Taken together, our study revealed that macrophages contribute to SCC expansion through interactions with tumor cells but are dispensable for SCC metastasis. Our study provides novel insights into understanding the contributions and limitations of TAMs in SCC progression.

Mouse Models for Studying Oral Cancer: Impact in the Era of Cancer Immunotherapy.

Model systems for oral cancer research have progressed from tumor epithelial cell cultures to in vivo systems that mimic oral cancer genetics, pathological characteristics, and tumor-stroma interactions of oral cancer patients. In the era of cancer immunotherapy, it is imperative to use model systems to test oral cancer prevention and therapeutic interventions in the presence of an immune system and to discover mechanisms of stromal contributions to oral cancer carcinogenesis. Here, we review in vivo mouse model systems commonly used for studying oral cancer and discuss the impact these models are having in advancing basic mechanisms, chemoprevention, and therapeutic intervention of oral cancer while highlighting recent discoveries concerning the role of immune cells in oral cancer. Improvements to in vivo model systems that highly recapitulate human oral cancer hold the key to identifying features of oral cancer initiation, progression, and invasion as well as molecular and cellular targets for prevention, therapeutic response, and immunotherapy development.

Isolation and propagation of keratinocytes derived from Cashmere goat fetus.

The study was conducted to isolate epidermal keratinocytes from Cashmere goat fetus with the aim to develop suitable conditions for keratinocyte cultivation and propagation. The methods developed for keratinocyte culture include (i) use of a feeder-layer of mitotically inactivated fibroblasts obtained from goat and mouse fetal skin, (ii) use of a substrate such as collagen IV, or (iii) without use of any substrate. Epidermal cell removal was established by enzymatically separating keratinocytes from 12 to 16 weeks aged fetal skin tissues treated with 0.125% trypsin solution overnight at 4 degrees C. The cells were maintained in all culture conditions with serum containing medium. Keratinocyte multiplication and proliferation were comparable in different culture conditions and the improved cellular attachment and growth have been obtained in cultures on feeder layers. Colony forming keratinocytes on feeder layer were heterogeneous in their growth potential. In feeder free conditions, high cellular density was required at plating for sub-cultivation as their poor attachment in culture dishes. This study reports the comparative efficacy of different culture conditions for keratinocyte isolation and in vitro propagation originating from Cashmere goat fetus.